The goal of this study is to utilize acoustic contrast agents with high-frequency ultrasound (HFU, >10 MHz) in order to image microcirculation and initiate gene transfection via sonoporation. Current contrast agents were not designed for these high frequencies. The study will assess two important applications: 1) imaging of slow-flow microvasculature in rabbit eyes and mouse embryos;and 2) the initiation of sonoporation in mouse embryos to assess the potential for gene transfection. Unlike with conventional imaging frequencies (<10 MHz), HFU provides the means to achieve fine-scale lateral resolution for microcirculation imaging and highly localized pressure exposure for sonoporation. Specifically, we propose undertaking comprehensive theoretical and experimental studies related to HFU excitation of acoustic contrast agents. Currently available protein-, lipid-, and polymer-shelled agents along with our own custom-made, polymer-shelled agents will be characterized experimentally to quantify backscatter (scattering cross section, nonlinear response, etc.), attenuation, speed of sound, and destruction threshold. Theoretical models that extend well-founded earlier models will be developed and simulations of radial response and agent destruction will be compared to experimental results. The models will be utilized to better understand the optimal material properties of contrast agents for HFU applications and to gain insight into the physical mechanisms that lead to the destruction of contrast agents. The theoretical and experimental results will then be used to formulate imaging and signal processing strategies to better visualize microcirculation. Knowledge gained from the initial experimental and theoretical work will also be applied to optimizing acoustic exposure conditions for in vivo animal experiments. The contrast agents will be used to image microcirculation in rabbit eyes and mouse embryos as well as to initiate gene transfection via sonoporation in mouse embryos. The animal experiments, particularly those involving the rabbit eye, will have direct relevance for imaging human diseases including glaucoma (2-3 million in U.S.), macular degeneration (200,000 new cases each year in U.S.), and primary intraocular tumors (3,000 new cases each year in U.S.). For instance, HFU with contrast agents may help diagnose glaucoma, the leading cause of blindness in the U.S., by permitting the assessment of microcirculation in the optic nerve and ciliary body. PUBLIC HEALTH RELEVANCE The ultimate goal of this research is to extend the use of acoustic contrast agents to higher frequencies in order to permit the imaging of microcirculation and the initiation of gene transfection via sonoporation. The knowledge we generate will assist in the design of new agents, optimizing acoustic exposure conditions, and, ultimately, will lead to new clinical applications such as ocular disease diagnosis and targeted gene transfection.